Food Analysis Introduction

FOOD ANALYSIS INTRODUCTION • Investigations in food science and technology, whether by the food industry, governmental agencies, or universities, often require determination of food composition and characteristics. • Trends and demands of consumers, the food industry, and national and international regulations challenge food scientists as they work to monitor food composition and to ensure the quality and safety of the food supply. • Food products require analysis as part of a quality management program throughout the development process (including raw ingredients), through production, and after a product is in the market. • Analysis is done of problem samples and competitor products Conti… • The characteristics of foods (i.e., chemical composition, physical properties, sensory properties) are used to answer specific questions for regulatory purposes and typical quality control. • The nature of the sample and the specific reason for the analysis commonly dictate the choice of analytical methods • Speed, precision, accuracy, and ruggedness often are key factors in this choice • Validation of the method for the specific food matrix being analyzed is necessary to ensure usefulness of the method • The success of any analytical method relies on the proper selection and preparation of the food sample, carefully performing the analysis, and doing the appropriate calculations and interpretation of the data. TRENDS AND DEMANDS: Food Industry • Many companies select suppliers, on whom they rely to perform the analytical tests to ensure compliance with detailed specifications for ingredients/raw materials • These specifications, and the associated tests, target various chemical, physical, and microbiological properties • Results of these analytical tests related to the predetermined specifications are delivered as a Certificate of Analysis (COA) with the ingredient/raw material • Companies must have in place a means to maintain control of these COAs and react to them. TRENDS AND DEMANDS: Government Regulations and International Standards and Policies • Government regulations and guidelines relevant to food analysis include • Nutrition labeling regulations, • Mandatory and voluntary standards • Good Manufacturing Practice (GMP) regulations (now called Current Good Manufacturing Practice in Manufacturing, Packing, or Holding Human Food), • Hazard Analysis Critical Control Point (HACCP) systems • The HACCP system is highly demanded of food companies by auditing firms and customers • The HACCP concept has been adopted not only by the US Food and Drug Administration (FDA) and other federal agencies in USA, but also by Codex Alimentarius Commission, an international organization that has become a major force in world food trade. TYPES OF SAMPLES ANALYZED • Chemical analysis of foods is an important part of a quality assurance program in • Selection of ingredients and raw materials • Food processing • Finished products • Chemical analysis also is important in • Formulating and developing new products • Evaluating new processes for making food products • Identifying the source of problems with unacceptable products STEPS IN ANALYSIS: Select and Prepare Sample • In analyzing food samples of the types described, all results depend on obtaining a representative sample and converting the sample to a form that can be analyzed • Neither of these is as easy as it sounds! • Sampling is the initial point for sample identification • Analytical laboratories must keep track of incoming samples and be able to store the analytical data from the analyses • This analytical information often is stored on a Laboratory Information Management System, or LIMS, which is a computer database program STEPS IN ANALYSIS: Perform the Assay • Performing the assay is unique for each component or characteristic to be analyzed and may be unique to a specific type of food product • It should be noted that numerous analytical methods utilize automated instrumentation, including auto samplers and robotics to speed the analyses STEPS IN ANALYSIS: Calculate and Interpret Results • To make decisions and take action based on the results obtained from performing the assay that determined the composition or characteristics of a food product, one must make the appropriate calculations to interpret the data correctly CHOICE AND VALIDITY OF METHOD: Characteristics of the Method • Numerous methods often are available to assay food samples for a specific characteristic or component • To select or modify methods used to determine the chemical composition and characteristics of foods, one must be familiar with the principles underlying the procedures and the critical step CHOICE AND VALIDITY OF METHOD: Objective of the Assay • Selection of a method depends largely on the objective of the measurement • For example, methods used for rapid online processing measurements may be less accurate than official methods used for nutritional labeling purposes CHOICE AND VALIDITY OF METHOD: Consideration of Food Composition and Characteristics • Proximate analysis of foods refers to determining the major components of moisture, ash (total minerals), lipids, protein, and carbohydrates • The performance of many analytical methods is affected by the food matrix (i.e., its major chemical components, especially lipid, protein, and carbohydrate) • In analysis, it is usually food matrix that presents the greatest challenge to the analyst • AOAC International, formerly known as the Association of Official Analytical Chemists (AOAC) • The AACC International, formerly known as the American Association of Cereal Chemists (AACC) CHOICE AND VALIDITY OF METHOD: Validity of the Method • A major consideration for determining method validity is the analysis of materials used as controls, often referred to as standard reference materials or check samples • Analyzing check samples concurrently with test samples is an important part of quality control • Besides government related groups, numerous organizations offer check sample services that provide test samples to evaluate the reliability of a method • The AACC International offers check samples such as flours, semolina, and other cereal-based samples, for analyses such as moisture, ash, protein, vitamins, minerals, sugars, sodium, total dietary fiber, soluble and insoluble dietary fiber, and β-glucan. Conti… • Samples also are available for testing physical properties and for microbiological and sanitation analyses • The American Oil Chemists’ Society (AOCS) has a reference sample program for oilseeds, oilseed meals, marine oils, aflatoxin, cholesterol, trace metals, specialty oils suitable for determination of trans fatty acids, and formulations for nutritional labeling • Laboratories from many countries participate in the program to check the accuracy of their work, their reagents, and their laboratory apparatus against the statistical norm derived from the group data AOAC INTERNATIONAL • AOAC International is an organization begun in 1884 to serve the analytical methods needs of government regulatory and research agencies • The goal of AOAC International is to provide methods that will be fit for their intended purpose (i.e., will perform with the necessary accuracy and precision under usual laboratory conditions). Sampling and Sample Preparation • Quality attributes in food products, raw materials, or ingredients are measurable characteristics that need monitoring to ensure that specifications are met • Some quality attributes can be measured online by using specially designed sensors and results obtained in real time (e.g., color of vegetable oil in an oil extraction plant) • However, in most cases quality attributes are measured on small portions of material that are taken periodically from continuous processes or on a certain number of small portions taken from a lot • The small portions taken for analysis are referred to as samples • The entire lot or the entire production for a certain period of time, in the case of continuous processes, is called a population • The process of taking samples from a population is called sampling • If the procedure is done correctly, the measurable characteristics obtained for the samples become a very accurate estimation of the population • A laboratory sample for analysis can be of any size or quantity SELECTION OF SAMPLING PROCEDURES General Information • The first step in any sampling procedure is to clearly define the population that is going to be sampled • The population may vary in size from a production lot, a day’s production, to the contents of a warehouse • Information obtained from a sample of a particular production lot in a warehouse must be used strictly to make inferences about that particular lot, but conclusions cannot be extended to other lots in the warehouse • Variance is an estimate of the uncertainty • The total variance of the whole testing procedure is equal to the sum of the variances associated with each step of the sampling procedure and represents the precision of the process • Precision is a measure of the reproducibility of the data • Accuracy is a measure of how close the data are to the true value • The most efficient way to improve accuracy is to improve the reliability of the step with the greatest variance that is frequently the initial sampling step • However, sample size is limited by time, cost, sampling methods, and the logistics of sample handling, analysis, and data processing Sampling Plan • Most sampling is done for a specific purpose, and the purpose may dictate the nature of the sampling approach • The two primary objectives of sampling are often • to estimate the average value of a characteristic • to determine if the average value meets the specifications defined in the sampling plan • Important categories of sampling purposes • 1. Nutritional labeling 2. Detection of contaminants and foreign matter 3. Statistical process control (Quality Assurance) 4. Acceptance of raw materials, ingredients, or products (Acceptance Sampling) 5. Release of lots of finished product 6. Detection of adulterations 7. Microbiological safety 8. Authenticity of food ingredients, etc • The International Union of Pure and Applied Chemistry (IUPAC) defines a sampling plan as: “A predetermined procedure for the selection, withdrawal, preservation, transportation, and preparation of the portions to be removed from a lot as samples” • A sampling plan should be a well-organized document that establishes the goals of the sampling plan, the factors to be measured, sampling point, sampling procedure, frequency, size, personnel, preservation of the samples, etc. • The primary aim of sampling is to obtain a sample, subject to constraints of size that will satisfy the sampling plan specifications Factors Affecting the Choice of Sampling Plans • The choice of a sampling plan is an important consideration, especially when monitoring food safety by measurement of fungal toxins, named mycotoxins, in food systems • Mycotoxins are distributed broadly and randomly within a population and a normal distribution cannot be assumed • sampling and good comminution and mixing prior to particle size reduction are more important than the chemical analysis itself Sampling by Attributes and Sampling by Variables • Sampling plans are designed for examination of either attributes or variables • Attribute sampling: sampling is performed to decide on the acceptability of a population based on whether the sample possesses a certain characteristic or not • Variable sampling: sampling is performed to estimate quantitatively the amount of a substance (e.g., protein content, moisture content, etc.) or a characteristic (e.g., color) on a continuous scale Acceptance Sampling • Acceptance sampling is a procedure that serves a very specific role: to determine if a shipment of products or ingredients has enough quality to be accepted • It can be performed by the food processor before receiving a lot of materials from a supplier, or by a buyer who is evaluating the processor’s output • Lot acceptance sampling plans that may be used for evaluation of attributes or variables, or a combination of both, fall into the following categories: single, double, multiple, sequential, and skip plans • In single sampling plans, the decision of accepting or rejecting a lot is based just on one sample of items taken at random • Double sampling plan (i.e., two samples are taken) • Multiple sampling plans are extensions of double sampling plans for which more than two samples are drawn to reach a conclusion • The ultimate extension of multiple sampling is sequential sampling • A sample is taken, and after analysis a decision of accepting, rejecting, or taking another sample is made • In skip lot sampling only a fraction of the submitted lots is inspected • It is a money-saving sampling procedure, but it can be implemented only when there is enough proof that the quality of the lots is consistent Risks Associated with Sampling • There are two types of risks associated with sampling: the producer’s and consumer’s risks • The consumer’s risk describes the probability of accepting a poor quality population • This should happen rarely (<5% of the lots), but the actual acceptable probability of a consumer risk depends on the consequences associated with accepting an unacceptable lot • The producer risk is the probability of rejecting (α) an acceptable product Homogeneous vs. Heterogeneous Populations • The ideal population would be uniform throughout and identical at all locations, Such a population would be homogeneous • Sampling from such a population is simple, as a sample can be taken from any location and the analytical data obtained will be representative of the whole • Even in an apparently uniform product, such as sugar syrup, suspended particles and sediments in a few places may render the population heterogeneous • Most populations that are sampled are heterogeneous • The location within a population where a sample is taken will affect the subsequent data obtained • However, sampling plans and sample preparation can make the sample representative of the population or take heterogeneity into account in some other way. Manual vs. Continuous Sampling • To obtain a manual sample the person taking the sample must attempt to take a “random sample” to avoid human bias in the sampling method • The sample must be taken from a number of locations within the population to ensure that it is representative of the whole population • For liquids in small containers, this can be done by shaking prior to sampling • Continuous sampling is performed mechanically. • An automatic sampling device is used to take liquid samples from a continuous production line • Continuous sampling should be less prone to human bias than manual sampling Probability Sampling • Probability sampling plans prescribe the selection of a sample from a population based on chance • It provides a statistically sound basis for obtaining representative samples with elimination of human bias • Simple random sampling requires that the number of units in the population be known, and each unit is assigned an identification number • Then using a random selection process, a certain number of identification numbers are selected according to the sample size • Systematic sampling is used when a complete list of sample units is not available, but when samples are distributed evenly over time or space, such as on a production line • The first unit is selected at random (random start) and then units are taken every nth unit (sampling interval) after that. Conti… • Composite sampling is used to obtain samples from bagged products such as flour, seeds, and larger items in bulk • Small aliquots are taken from different bags, or containers, and combined in a simple sample (the composite sample) that is used for analysis. Problems in Sampling • Errors also may be introduced by not understanding the population distribution and subsequent selection of an inappropriate sampling plan. • Unreliable data also can be obtained by nonstatistical factors such as poor sample storage resulting in sample degradation • Refrigeration or freezing may be necessary to protect chemically unstable samples • However, freezing should be avoided when storing unstable emulsions. • Preservatives (e.g., mercuric chloride, potassium dichromate, and chloroform) can be used to stabilize certain food substances during storage. Mislabeling and official samples • Mislabeling of samples causes mistaken sample identification • Samples should be clearly identified by markings on the sample container in a manner such that markings will not be removed or damaged during storage and transport • For example, plastic bags that are to be stored in ice water should be marked with water-insoluble ink • If the sample is an official or legal sample the container must be sealed to protect against tampering and the seal mark easily identified • Official samples also must include the date of sampling with the name and signature of the sampling agent. The chain of custody of such samples must be identified clearly. PREPARATION OF SAMPLES General Size Reduction Considerations • If the particle size or mass of the sample is too large for analysis, it must be reduced in bulk or particle size • To obtain a smaller quantity for analysis the sample can be spread on a clean surface and divided into quarters • The two opposite quarters are combined • If the mass is still too large for analysis, the process is repeated until an appropriate amount is obtained • This method can be modified for homogeneous liquids by pouring into four containers, and it can be automated • The samples are thus homogenized to ensure negligible differences between each portion Conti.. • In case of meat and meat products, it is specified that small samples should be avoided, as this results in significant moisture loss during preparation and subsequent handling • A further example of size reduction is the preparation of solid sugar products for analysis as described in AOAC Method. • The method prescribes that the sugar should be ground, if necessary, and mixed to uniformity • Raw sugars should be mixed thoroughly and rapidly with a spatula • Lumps are to be broken by a mortar and pestle or by crushing with a glass or iron rolling pin on a glass plate. Grinding • Grinding is important for sample preparation prior to analysis and for food ingredient processing • Various mills are available for reducing particle size to achieve sample homogenization • To homogenize moist samples, bowl cutters, meat mincers, tissue grinders, mortars and pestles, or blenders are used, while mortars and pestles and mills are best for dry samples • Some foods are more easily ground after drying in a desiccator or vacuum oven • Grinding wet samples may cause significant losses of moisture and chemical changes • In contrast, grinding frozen samples reduces undesirable changes • The grinding process should not heat the sample, and therefore the grinder should not be overloaded because heat will be produced through friction • Contact of food with bare metal surfaces should be avoided if trace metal analysis is to perform Preventing Changes in Sample Once we have selected our sample, we have to ensure that: • It does not undergo any significant changes in its properties from the moment of sampling to the time when the actual analysis is carried out, e.g., enzymatic, chemical, microbial or physical changes. 35 There are a number of ways these changes can be prevented Enzymatic Inactivation • Many foods contain active enzymes they can cause changes in the properties of the food prior to analysis, e.g., proteases, cellulases, lipases, etc. • If the action of one of these enzymes alters the characteristics of the compound being analyzed then it will lead to erroneous data and it should therefore be inactivated or eliminated • { Freezing, drying, heat treatment and chemical preservatives} 36 There are a number of ways these changes can be prevented Lipid Protection • Unsaturated lipids may be altered by various oxidation reactions • Exposure to light, elevated temperatures, oxygen or pro-oxidants can increase the rate at which these reactions proceed • Consequently, it is usually necessary to store samples that have high unsaturated lipid contents under nitrogen or some other inert gas, in dark rooms or covered bottles and in refrigerated temperatures. • Therefore, ideally unsaturated lipids should be extracted just prior to analysis 37 There are a number of ways these changes can be prevented Microbial Growth and Contamination • Microorganisms are present naturally in many foods and if they are not controlled. they can alter the composition of the sample to be analyzed. • Freezing, drying, heat treatment and chemical preservatives are often used to control the growth of microbes in foods. • Physical Changes. A number of physical changes may occur in a sample, e.g., water may be lost or gained due to condensation; fat crystallize; structural properties may be disturbed. • Physical changes can be minimized by controlling the temperature of the sample, and the forces that it experiences 38 COMPOSITIONAL ANALYSIS OF FOOD Moisture and Total Solids Importance of Moisture Assay • Moisture assays can be one of the most important analyses performed on a food product and yet one of the most difficult from which to obtain accurate and precise data • The dry matter that remains after moisture removal is commonly referred to as total solids • This analytical value is of great economic importance to a food manufacturer because water is an inexpensive filler • The listing gives some examples in which moisture content is important to the food processor 40 Conti… 1. Moisture is a quality factor in the preservation of some products and affects stability in (a) Dehydrated vegetables and fruits (b) Dried milks (c) Powdered eggs (d) Dehydrated potatoes (e) Spices and herbs 2. Moisture is used as a quality factor for (a) Jams and jellies to prevent sugar crystallization (b) Sugar syrups (c) Prepared cereals – conventional, 4–8%; puffed, 7–8% 42 Conti… 3. Reduced moisture is used for convenience in packaging or shipping of (a) Concentrated milks (b) Liquid cane sugar (67% solids) and liquid corn sweetener (80% solids) (c) Dehydrated products (these are difficult to package if too high in moisture) (d) Concentrated fruit juices 4. Moisture (or solids) content is often specified in compositional standards (i.e., Standards of Identity) (a) Cheddar cheese must be ≤39% moisture (b) Enriched flour must be ≤15% moisture (c) Pineapple juice must have soluble solids of ≥10.5˚Brix (conditions specified). 43 Conti… (d) Glucose syrup must have ≥70% total solids. (e) The percentage of added water in processed meats is commonly specified. 5. Computations of the nutritional value of foods require that you know the moisture content. 6. Moisture data are used to express results of other analytical determinations on a uniform basis [i.e., dry weight basis (dwb), rather than wet weight basis (wwb)] Moisture Content of Foods • The approximate, expected moisture content of a food can affect the choice of the method of measurement • It can also guide the analyst in determining the practical level of accuracy required when measuring moisture content, relative to other food constituents. 44 Forms of Water in Foods • The ease of water removal from foods depends on how it exists in the food product. The three states of water in food products are: • Free water: This water retains its physical properties and thus acts as the dispersing agent for colloids and the solvent for salts • Adsorbed water: This water is held tightly or is occluded in cell walls or protoplasm and is held tightly to proteins • Water of hydration: This water is bound chemically, for example, lactose monohydrate; also, some salts such as Na2SO4·10H2O • Depending on the form of the water present in a food, the method used for determining moisture may measure more or less of the moisture present • This is the reason for official methods with stated procedure 45 Sample Collection and Handling • Procedures are perhaps the greatest potential source of error in any analysis • Precautions must be taken to minimize inadvertent moisture losses or gains that occur during these steps • Any exposure of a sample to the open atmosphere should be as short as possible • Any heating of a sample by friction during grinding should be minimized • Headspace in the sample storage container should be minimal because moisture is lost from the sample to equilibrate the container environment • Control temperature fluctuations since moisture will migrate in a sample to the colder part 46 OVEN DRYING METHODS • Any oven method used to evaporate moisture has its foundation, the fact that the boiling point of water is 100◦C; however, this considers only pure water at sea level • Free water is the easiest of the three forms of water to remove • Moisture removal is sometimes best achieved in a two-stage process • Liquid products (e.g., juices, milk) are commonly pre-dried over a steam bath before drying in an oven • Products such as bread and field-dried grain are often air dried, then ground and oven dried, with the moisture content calculated from moisture loss at both air and oven drying steps 47 Conti… • Particle size, particle size distribution, sample sizes, and surface area during drying influence the rate and efficiency of moisture removal. Decomposition of Other Food Constituents • Moisture loss from a sample during analysis is a function of time and temperature • Decomposition enters picture when time is extended too much, or temperature is too high • Thus, most methods for food moisture analysis involve a compromise between time and a particular temperature at which limited decomposition might be a factor • One major problem exists in that the physical process must separate all the moisture without decomposing any of the constituents that could release water 48 Conti… • The moisture generated in carbohydrate decomposition is not the moisture that we want to measure. • Certain other chemical reactions (e.g., sucrose hydrolysis) can result in utilization of moisture, which would reduce the moisture for measurement • A less serious problem, but one that would be a consistent error, is the loss of volatile constituents, such as acetic, propionic, and butyric acids; and alcohols, esters, and aldehydes among flavor compounds 49 Temperature Control • Determine extent of variation within oven before relying on data collected from its use. • Consider the temperature variation in three types of ovens: convection (atmospheric), forced draft, and vacuum • The greatest temperature variation exists in a convection oven • This is because hot air slowly circulates without the aid of a fan. • A convection oven should not be used when precise and accurate measurements are needed. • Forced draft ovens have the least temperature differential across the interior of all ovens, usually not greater than 1◦C • Air is circulated by a fan that forces air movement throughout the oven cavity 50 Types of Pans for Oven Drying Methods • Pans used for moisture determinations are varied in shape and may or may not have a cover • The AOAC International moisture pan is about 5.5 cm in diameter with an insert cover • Pan covers are necessary to control loss of sample by spattering during the heating process • If the cover is metal, it must be slipped to one side during drying to allow for moisture evaporation 51 Handling and Preparation of Pans • The preparation and handling of pans before use requires consideration • Use only tongs to handle any pan, even fingerprints have weight • All pans must be oven treated to prepare them for use • This is a factor of major importance unless disproved by the technologist doing moisture determinations with a particular type of pan • Disposable aluminum pans must be vacuum oven dried for 3 h before use • Store dried moisture pans in a functioning desiccator • The glass fiber covers should be dried for 1 h before use 52 Control of Surface Crust Formation (Sand Pan Technique) • Some food materials tend to form a semipermeable crust or lump together during drying, which will contribute to erratic and erroneous results • To control problem, analysts use the sand pan technique • Clean, dry sand and a short glass stirring rod are pre-weighed into a moisture pan • Subsequently, after weighing in a sample, the sand and sample are admixed with the stirring rod left in the pan • The remainder of the procedure follows a standardized method if available; otherwise the sample is dried to constant weight • The purpose of the sand is twofold: to prevent surface crust from forming and to disperse the sample so evaporation of moisture is less impeded 53 Calculations • Moisture and total solids contents of foods can be calculated as follows using oven drying procedures 54 Forced Draft Oven • When using a forced draft oven, the sample is rapidly weighed into a pre-dried moisture pan covered and placed in the oven for an arbitrarily selected time if no standardized method exists • Drying time periods for this method are 0.75–24 h, depending on the food sample and its pretreatment; some liquid samples are dried initially on a steam bath at 100˚C to minimize spattering 55 Vacuum Oven • By drying under reduced pressure (25–100 mm Hg), one is able to obtain a more complete removal of water and volatiles without decomposition within a 3–6-h drying time • Vacuum ovens need a dry air purge in addition to temperature and vacuum controls to operate within method definition 56 Microwave Analyzer • Determination of moisture using a standard oven, which, though accurate, can take many hours to dry a sample • Other methods have been developed over the years including infrared and various types of instruments that utilize halogen lamps or ceramic heating elements • They were often used for “spot checking” because of their speed, but they lacked the accuracy of the standard oven method • The introduction of microwave moisture/solids analyzers gave laboratories the accuracy and the speed they wanted • Microwave moisture analysis, often called microwave drying, was the first precise and rapid technique that allowed some segments of the food industry to make in-process adjustment of the moisture content in food products before final packaging 57 Infrared Drying • Infrared drying involves penetration of heat into the sample being dried, as compared with heat conductivity and convection with conventional ovens • Such heat penetration to evaporate moisture from the sample can significantly shorten the required drying time to 10–25 min • The analyst must be careful that the sample does not burn or case harden while drying • Infrared drying ovens may be equipped with forced ventilation to remove moisture air and an analytical balance to read moisture content directly • No infrared drying moisture analysis techniques are approved by AOAC International currently • However, because of the speed of analysis, this technique is suited for qualitative in- process use 58 Rapid Moisture Analyzer Technology • Compact instruments are available, such as analyzers that detect moisture levels from 50 ppm to 100% using sample weights of 150 mg to 40 g (e.g., Computrac, Arizona Instrument LLC, Chandler, AZ) • Using a digital balance, the test sample is placed on an aluminum pan or filter paper and the heat control program (with a heating range of 25–275◦C) elevates the test sample to a constant temperature • As the moisture is driven from the sample, the instrument automatically weighs and calculates the percentage moisture or solids • These analyzers are utilized for both production and laboratory use with results comparable to reference methods 59